1. Field of the Invention
The present invention relates to an optical bioimaging technology for biological specimens such as small animals. More particularly, the present invention relates to a technique for avoiding the degradation of images of a living body simultaneously picked up by a multi-directional image acquisition device due to an illumination backlight problem.
2. Description of the Related Art
In medical and biological fields, imaging of the distribution of molecular species in a living body is an important research technique. Heretofore, cellular-level imaging of molecular species has been widely performed using a microscope and a molecular probe labeled with fluorochrome or a chemiluminescent molecular probe. On the other hand, there has been a demand for a device capable of observing the distribution of molecular species of interest in a living body not at the cellular level but at the level of organ or the entire animal body larger than cells. Such a device makes it possible to acquire images of a living body, such as a mouse, whose cancer cells are labeled with fluorescence probes every day or every week to monitor the growth of cancer cells of interest with the lapse of time. In a case where the growth of cancer cells in the body of an animal is observed by a conventional cellular-level observation device, it is necessary to kill the animal to stain a site to be observed or to attach a fluorescent substance to cancer cells. In this case, however, it is impossible to monitor the growth of cancer cells in one animal body over a long period of time. For this reason, there has been a demand for development of a device capable of observing molecular species in the body of a living small animal to obtain internal information of the small animal.
Near-infrared light relatively easily penetrates a living body. Therefore, light ranging from about 600 nm to 900 nm is used in devices for observing small animals. However, according to a conventional observation technique, a specimen is usually observed only from above and cannot be simultaneously observed from various directions. Therefore, there is a case where, for example, when a mouse is observed from a specific direction, cancer is not detected, but when the mouse is observed from a direction opposite to the specific direction, cancer is detected. When a mouse is observed using an unidirectional observation device, an operator has no choice to observe the mouse by a method approximate to multi-directional observation by picking up multi-directional images of the mouse rotated about its body axis by small increments. However, in this case, reproducible data cannot be obtained, and the mouse cannot be simultaneously observed from various directions. Particularly, in the case of observation of luminescence from a living body, the intensity of luminescence is very weak, and therefore, it is usually necessary to perform integrated exposure on a two-dimensional detector for several tens of seconds to a few minutes. On the other hand, the intensity of luminescence changes with time, and therefore, when image pickup is performed every time the observation direction is changed, image pickup conditions are different among image pickup directions and, thus, resulting images are useless. For this reason, it is preferred that two or more images of a living body picked up from two or more directions can be simultaneously and parallelly integrated on a detector for a long period of time. In the case of fluorescence observation, it is possible to perform fluorescence image measurement in a relatively short period of time. Still, it is absolutely necessary to simultaneously pick up information from various directions to speedily obtain accurate data.
As a method for acquiring multi-directional images, one for sequentially acquiring images observed from various angles using a rotating reflection mirror in a time-sharing manner is known (see Patent Document 1). According to this method, a specimen can be observed from various directions by rotating the mirror and by changing the position of the specimen itself by parallel displacement, and therefore, it is not necessary to rotate the specimen and a two-dimensional detector. However, the method disclosed in Patent Document 1 uses a rotating reflection mirror and therefore has the following drawbacks: a specimen is measured from various directions in a time-sharing manner, that is, multi-directional simultaneous measurement cannot be performed, and therefore, it takes a long time to complete measurement; images observed from different directions are picked up at different times and therefore lack in accuracy in the case of, for example, luminescence measurement because the intensity of luminescence changes with time; and a device using a rotating reflection mirror has a complicated structure.
Some image pickup optical systems not using a rotating reflection mirror are also known (see Patent Documents 2 to 9). Patent Documents 2 and 3 each describe the concepts of multi-directional observation and an image pickup optical system for multi-directional observation including a reflection mirror and a two-dimensional detector. The multi-directional observation devices described in Patent Documents 2 and 3 are merely target devices of the present invention, and there is no description about a backlight problem in Patent Documents 2 and 3. Patent Document 4 relates to a technique for “utilizing” light from the opposite side of a camera, and therefore has nothing in common with the present invention dealing with a technique for “avoiding” light from the opposite side of a camera. Patent Document 6 does not describe a technique for multi-directional observation of a specimen but describes a technique for picking up a panoramic (360°) image around a camera. Therefore, Patent Document 6 has no bearing on the present invention because the relationship between a specimen and a camera is the reverse of that in the present invention. Patent Documents 5, 7, and 8 do not describe even multi-directional observation. Patent Document 7 shows concentric arrangement of light sources in FIG. 1 by chance, but this arrangement is not intended to overcome a backlight problem and therefore gives no clue to the solution of problems to be solved by the present invention. Each of the Patent Documents 2 to 9 will be further described in detail as follows.
Patent Document 2 relates to an invention for photographing a three-dimensional image of an object. The only similarity between this invention and the present invention is the use of a polygonal mirror, but this invention is not intended to acquire biological information such as fluorescence images. Further, Patent Document 2 describes that the angle of the reflection mirror is made variable to increase the number of observation directions, but this means that observation is performed in a partial time-sharing manner, whereas the present invention is intended to perform simultaneous observation. Further, there is a description about the use of a light source, but this light source is arranged simply to illuminate a dark back side of a specimen. Therefore, Patent Document 2 does not, of course, describe a backlight problem caused by the light source for illuminating the back side of a specimen, and does not at all consider the possibility of direct entry of light from the light source into a camera. For this reason, the invention disclosed in Patent Document 2 does not provide any clue to the solution of a backlight problem associated with multi-directional simultaneous measurement which is a problem to be solved by the present invention.
Patent Document 3 proposes the use of a cylindrical convex mirror for expanding the image pickup range of a video camera placed along a conveyor for transporting fruit. This invention is similar to the present invention in that an image of an object reflected by a reflection mirror as well as a real image of the object is picked up. However, Patent Document 3 does not describe even an irradiation light source, and therefore does not give any clue to the solution of a backlight problem associated with multi-directional simultaneous measurement which is a problem to be solved by the present invention.
Patent Document 4 relates to a device for detecting defective parts by detecting leak light from an inspection surface of a printed-wiring board or the like, and describes a method for detecting light leaking through a printed-wiring board to be inspected when a light source is placed on the back side of the printed-wiring board. As described above, Patent Document 4 describes a technique for “utilizing” light from the opposite side of a camera, whereas the present invention deals with a technique for “avoiding” backlight from the opposite side of a camera. Therefore, the invention disclosed in Patent Document 4 has nothing in common with the present invention dealing with a technique for “avoiding” light from the opposite side of a camera. In this invention, light reflected by a defective part happens to be received by two or more mirrors arranged in different directions, but this does not give any clue to the solution of a backlight problem associated with multi-directional simultaneous measurement which is a problem to be solved by the present invention.
Patent Document 5 relates to a device for detecting a fluorescence image of a biological specimen having been subjected to, for example, electrophoresis by a CCD camera. When the size of a specimen is changed, the specimen needs to be moved toward or away from the camera. Therefore, a technique for compensating for the fluctuation of data caused by the difference in distance from a fluorescence excitation light source is described in Patent Document 5. In the case of this device, however, fluorescence is observed from only one direction, and in addition, the disadvantage of backlight is not perceived as a problem at all, although two light sources for irradiating a specimen with excitation light from two directions, that is, a bottom excitation light source 51 and a downward irradiating excitation light source 52 are shown in FIG. 2 in Patent Document 5. Therefore, although Patent Document 5 describes a fluorescence detection device, there is no description that gives a clue to the solution of problems to be solved by the present invention.
Patent Document 6 deals with an invention relating to a panoramic camera, that is, a technique for picking up a panoramic (360°) image around a camera. On the other hand, the present invention is directed to a multi-directional measurement device for picking up images of a specimen (object to be measured) placed at the center from various directions around the specimen. Considering that the positional relationship between a specimen and a camera is the reverse of that in the present invention, it is self-evident that Patent Document 6 has no bearing on the present invention. Although the camera disclosed in Patent Document 6 happens to use a polygonal reflection mirror for picking up a panoramic (360°) image around it, Patent Document 6 has nothing to do with a backlight problem to be solved by the present invention.
Patent Document 7 relates to a technique to be used for a device, such as a security camera, for picking up images of an object from one direction by simultaneously irradiating the object with light from three or more directions. More specifically, the frequencies (i.e., wavelengths) of irradiation light are made different among irradiation directions, and in addition, the filter of each pixel of a detector provided in a camera is selected so as to transmit any one of frequencies (wavelengths) different among irradiation directions. This makes it possible to acquire three or more images of an object different in irradiation direction while the object is simultaneously irradiated from three or more directions. The obtained images different in irradiation direction are weighted. There is some relation between Patent Document 7 and the present invention in multi-directional “illumination”. However, Patent Document 7 discloses a unidirectional image pickup device and does not describe a backlight problem at all, whereas the present invention deals with a problem associated with multi-directional image pickup. In FIG. 1 of Patent Document 7, a camera happens to be pointed toward a light source, but it is not perceived as a problem in Patent Document 7. Therefore, it cannot be said that Patent Document 7 deals with a backlight problem, although the number of observation directions is only one. For this reason, it can be hardly said that the description of Patent Document 7 gives a clue to the solution of a backlight problem associated with multi-directional observation.
Patent Document 8 describes an invention relating to an endoscope. In Patent Document 8, image pickup using irradiation light having different wavelengths and a fluorescence image pickup device are described, and therefore, a fluorescence excitation light source is of course described. However, there is no description about multi-directional observation and a backlight problem caused by excitation light emitted from the fluorescence excitation light source. For this reason, Patent Document 8 does not at all give a clue to the solution of a backlight problem associated with multi-directional observation which is a problem to be solved by the present invention.
Patent Document 9 discloses backside illumination during fluorescence measurement. More specifically, a fluorescence excitation light source is arranged so as to be brought into contact with the back surface of a biological specimen so that the light source cannot be seen from a camera. That is, the light source is hidden by a specimen itself. This technique enables backside illumination in unidirectional observation, but definitely cannot be used in multi-directional observation. Further, strong light enters a detector when a specimen is removed. Therefore, the invention disclosed in Patent Document 9 is absolutely different from the present invention because the present invention is directed to a structure capable of preventing direct light from a light source from entering a detector regardless of the presence or absence of a specimen.    Patent Document 1: US Patent Application Publication No. 20050201614    Patent Document 2: JP-A-2001-330915    Patent Document 3: JP-A-10-124648    Patent Document 4: JP-A-3-111745    Patent Document 5: JP-A-2003-287497    Patent Document 6: JP-A-8-125835    Patent Document 7: WO 2007/108070    Patent Document 8: JP-A-2005-13611    Patent Document 9: U.S. Pat. No. 6,922,246